Molecular marker and application of agpl1 micro-controlling gene of rice amylose content
A technology of amylose content and molecular markers, applied in the field of agricultural biotechnology engineering, can solve the problems of poor accuracy and achieve the effect of overcoming the long time period
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Embodiment 1
[0063] Example 1. Using the Indel marker AGPL1-m to identify polymorphisms in the japonica rice Nipponbare with low amylose content and the indica rice Teqing with high amylose content
[0064] The specific method is: select rice materials Nipponbare and Teqing from the germplasm resource bank of the China Rice Research Institute, and use Nipponbare and Teqing to obtain their F1, and use the primer AGPL1-m to identify their polymorphism ( figure 1 ).
[0065] 1. DNA extraction
[0066] 1) Prepare DNA extraction buffer:
[0067] Add 1 volume of DNA extraction solution (0.35M sorbitol; 0.1M Tris, pH8.2; 0.005MEDTA; the rest is water), 1 volume of nuclear lysis solution (0.2M Tris, pH7.5; 0.05M EDTA; 2M NaCl; 0.055MCTAB; the rest is water) and 0.4 volume of 5% (mass concentration) sarkosyl solution (i.e. the aqueous solution of sodium lauryl-N-methylglycinate); finally add sodium bisulfite to prepare DNA extraction buffer solution; the final concentration of sodium bisulfite i...
Embodiment 2
[0087] Example 2. Using the Indel molecular marker AGPL1-m to identify sequence differences between the japonica rice Nipponbare with low amylose content and the indica rice Teqing with high amylose content
[0088] The specific method is: use the Indel molecular marker AGPL1-m to perform PCR amplification on the genomic DNA of Nipponbare and Teqing, entrust Shanghai Yingjun Biotechnology Co., Ltd. to sequence the amplified products, and compare the differences in their sequences ( figure 2 ).
[0089] 1. DNA extraction
[0090] 1) Prepare DNA extraction buffer:
[0091] With embodiment 1.
[0092] 2), the paddy rice blade of above-mentioned Nipponbare and special green is carried out as follows respectively:
[0093] With embodiment 1.
[0094] 3) PCR amplification
[0095] With embodiment 1.
[0096] 4) Recovery of PCR products
[0097] The recovery of PCR products was carried out by using the PCR product recovery kit (spin column type, catalog number: DP1403) develo...
Embodiment 3
[0099] Example 3. Using the Indel marker AGPL1-m to carry out assisted selection breeding for low amylose content
[0100] The specific method is: the gene donor parent Nipponbare with low amylose content is crossed, backcrossed and self-crossed with Teqing, an indica rice variety with high amylose content, and the segregation population is selected by the assisted selection of the resulting offspring combined with the molecular marker AGPL1-m The individual plants with the same middle zone type and the Japanese sunny zone type are used for breeding and improvement.
[0101] 1. DNA extraction
[0102] 1) Prepare DNA extraction buffer:
[0103] With embodiment 1.
[0104] 2), above-mentioned Nipponbare, Teqing, and the rice blades of the gained offspring are respectively processed as follows:
[0105] With embodiment 1.
[0106] 2. Indel marker detection
[0107] 1), PCR amplification
[0108] With embodiment 1.
[0109] 2), electrophoresis detection
[0110] With embod...
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